The present invention relates to a sustained release and long residing ophthalmic formulation and the process of preparing the same.
Medication of the eyes is done for two purposesxe2x80x94to treat the outside of the eyes for such infections as conjunctivitis, blepharitis, keratitis sicca etc and to provide intraocular treatment through the cornea for diseases such as glaucoma or uveitis. Most ocular diseases are treated with topical application of solutions administered as eye drops. One of the major problems encountered with the topical delivery of ophthalmic drugs is the rapid and extensive precorneal loss caused by drainage and high tear fluid turn over. After instillation of an eye-drop, typically less than 2-3% of the applied drug penetrates the cornea and reaches the intra-ocular tissue, while a major fraction of the instilled dose is often absorbed systematically via the conjunctiva and nasolacrimal duct. Another limitation is relatively impermeable corneal barrier that limits ocular absorption.
Because of the inherent problems associated with the conventional eye-drops there is a significant efforts directed towards new drug delivery systems for ophthalmic administration such as hydrogels, micro- and nanoparticles, liposomes and collagen shields. Ocular drug delivery is an approach to controlling and ultimately optimizing delivery of the drug to its target tissue in the eye. Most of the formulation efforts aim at maximizing ocular drug absorption through prolongation of the drug residence time in the cornea and conjunctival sac as well as to slow drug release from the delivery system and minimizing precorneal drug loss.
To solve the above mentioned problem associated with the ocular delivery of drugs. WO 9405257 A1 940317 discloses a method for preparing a bioerodible drug delivery vehicle composed of solid polymeric matrix formed from derivatised cellulose and methacrylic acid copolymer and incorporating ophthalmic drugs in it. The inventors have demonstrated that such formulation when instilled in eyes the polymeric materials bio-erodes and dispenses the incorporated drug on the cornea surface. However, the problem always associated with the use of such bulk gel is the blurring effect and bio-compatibility of the polymeric material. Moreover, the long residence time and sustained release of drug on cornea surface have not been achieved.
There have been reported other studies on the use of co-polymeric materials as earners for ophthalmic drugs and particularly noteworthy are the attempts to incorporate hydrophobic drugs into the hydrophobic core of the copolymer micelles. The pharmaceutical efficacy of these formulations depends on the specific nature and properties of the co-polymeric materials.
For example, EP 0744938 A1 961204 discloses sustained release liquid aqueous ophthalmic delivery system. The method provides a slow and sustained release of treating agents. The polymer used was chitosan and was applied in the form of bulk material. However, it is difficult for such bulk polymer to penetrate the conceal membrane and the liquid formulation remain in the liquid form even at body temperature so that there exists every possibility to be washed away by tears. There also exists a problem of this formulation. The acidic pH of the liquid pH 3.0 to 6.2, is not very much patient compliance. The other patents with related formulation with chitosan is WO 9522315 A1 950824 and EP 0594760.
A1 940504 discloses a method for entrapping ophthalmic drugs in polyacryl acid polymer to obtain a fluid aqueous gel having a pH of between 6.5 and 8. The formulation is claimed to be useful for treating various pathological ocular conditions. This method, thus solves the potential muco-adhesive problem of the polymer by using poly-acrylic acid gels. However, pure polyacryl acid is so much sticky that a permanent polymer layer on the cornea surface may cause blurred vision. Moreover, being bulk gel, these polymeric composition may not have sufficient penetration and subsequently less bio-availability. The same is the comment for other patents: WO 9300887 A1 930121 and WO 9922713 A1 19990514 and WO 9405257 A1 940317.
There are reported studies on formulations of non-steroidal anti-inflammatory drugs (NSAID) using different carriers particularly directed towards oral delivery to avoid the adverse effects on the gastrointestinal tract.
A patent WO 93/25190 has disclosed surface modified NSAID nanoparticles by taking crystalline NSAID having surface modifier adsorbed on the surface thereof in an amount sufficient to maintain an effective average particle size of less than about 400 nm.
One patent, EP 0274870 A2 880720, has disclosed the micelles containing polyethoxylated nonionic surfactant as carrier for certain NSAIDs. Other patents. EP 0818992 A1 980121 and WO 9505166 A1 950223 disclose the similar type of micellar encapsulation of NSAIDs using cellular acetate phthalate and gelatin.
Another patent, WO 9702017 A1 970123 used hydrophilic poloxamer for encapsulating NSAIDs. These are some of the patents, which are mainly directed towards oral delivery formulations so that these NSAIDs cannot be exposed in GI tract. These formulations do not have any relevance to eye drop preparations.
To overcome the problem of blurred vision and poor bio-availability of drug by using bulk gel in ophthalmic formulations, it has been suggested that colloidal carriers would have better effect. Colloidal carriers which have been studied for ocular delivery are mainly liposomes and nanoparticles because of their extremely small size. The main limitation of liposomes as ocular drug delivery system is its surface charge. Positively charged liposomes seemed to be preferentially captured at the negatively charged corneal surface compared to neutral and negatively charged liposomes. Another limitation of liposomes is the instability of the lipid aggregates on the mucine surface. The vesicular aggregates of positively charged lipids are completely disintegrated on the negatively charged mucine membrane surface.
Nanoparticles as drug carriers for ocular delivery have been revealed to be more efficient than liposomes and in addition to all positive points of liposomes, these nanoparticles are exceptionally stable entity and the sustained release of drug can be modulated.
Some studies have been reported in the literature as well as patents on the use of nanoparticles carriers for encapsulating water insoluble drugs. For example, a U.S. Pat. No. 5,510,103 disclose the entrapment of water-insoluble drugs including NSAIDs in the hydrophobic core of polymeric micelles of different block copolymers. Another U.S. Pat. No. 5,449,513 has also disclosed the use of various amphiphilic copolymers in the form of micelles to physically entrap water-insoluble drugs. The other patents of similar type are U.S. Pat. No. 5,510,103, U.S. Pat. No. 5,449,513, U.S. Pat. No. 5,124,151.
All these patents described the method of preparation of amphiphilic copolymers of random compositions of two or three different types of amphiphilic monomers and aggregating them in aqueous solutions and dissolving the water-insoluble drugs inside the hydrophobic core of these polymeric micelles. The properties of these copolymers are governed by the composition of them. But none of the polymers could comply the overall requirements of best ophthalmic formulationxe2x80x94hydrogel formation, mucoadhesiveness, thermosensitivity and small particle sizexe2x80x94in a single formulation.
The object of the present invention is to obviate the above-mentioned drawbacks and to provide a formulation having thermosensitivity, mucoadhesiveness, hydrogel properties and small particle size all together in a single composition.
Still further object of this invention is to provide sustained release and long residing ophthalmic formulation, so that the release of the entrapped drug can be controlled and the process of preparing the same.
To achieve the said objectives, this invention provides a sustained release and long residing ophthalmic formulation comprising:
micelle solution of block random co-polymer having a hydrophobic component and a hydrophilic component of general formula xe2x80x94(X+Y+Z-)m, wherein
m is an integer greater than 2,
X is a monomer which will provide hydrogel formation properties of the co-polymer to reduce the irritability of the eye and is selected from vinyl group of compounds,
Y is a monomer which will provide thermosensitivity properties of the co-polymer having a general formula R1xe2x80x94R2Nxe2x80x94(Cxe2x95x90O)xe2x80x94CHxe2x95x90CH2, where R1=a proton or CnH2n+1 in which n may have the value from 3 to 6 and R2=alkyl group having chain length of C3 to C6 
Z is a monomer which will provide mucoadhesiveness and pH-sensitivity properties to the co-polymer and is selected from acrylate based monomers.
and
at least one hydrophobic drug with the said block co-polymer solution;
The said vinyl group of compounds is selected from vinyl pyrrolidone, vinyl alcohol, vinyl chloride or vinyl acetate.
Y is N-isopropyl acrylamide(NIPAAM)
The said acrylate based monomers is selected from acrylic acid or methyl methacrylate.
xe2x80x98mxe2x80x99 lies in the range 10-4000.
The said drug is a hydrophobic ohphthalmic drug and is selected from the family of non steroidal anti inflammatory drug (NSAID).
The said NSAID drug is selected from 5-benzoyl (2,3 dihydro-1H pyrrolizone-1carboxylic acid); 1-(4 chlorobenzoyl)-5 methoxy-2-methyl-1H-indole-3acetic acid or N-(4-nitro-2 phenoxyphenyl) methanesulphonamide or a mixture of at least any two thereof.
The said drug used for entrapment is in solution or in powder form.
The said drug is hydrophobic.
The said polymerized micelles containing the hydrophobic drug have a size in the range of 10 nm-100 nm diameter
The said polymerized micelles containing the hydrophobic drug have a size in the range of 10 nm to 50 nm.
The random block copolymers, made of amphiphilic monomers, are biocompatible and non-antigenic materials.
The optimum molar ratio of the combination of amphiphilic monomers X, Y and Z is 9.5%:85.7%:4.8% respectively from the point of view of mucoadhesiveness and thermosensitivity of the co-polymer.
The present invention further provides a process for preparing a sustained release and long residing ophthalmic formulation comprising the steps of:
preparing micelle solution of block random co-polymer having a hydrophobic component and a hydrophilic component of general formula xe2x80x94(X+Y+Z-)m, wherein
m is an integer greater than 2
X is a monomer which will provide hydrogel formation properties of the co-polymer to reduce the irritability of the eye and is selected from vinyl group of compounds
Y is a monomer which will provide thermosensitivity properties of the co-polymer having a general formula R1xe2x80x94R2Nxe2x80x94(Cxe2x95x90O)xe2x80x94CHxe2x95x90CH2, R1=a proton or CnH2n+1 in which n may have the value from 3 to 6 and R2=alkyl group having chain length of C3 to C6 
Z is a monomer which will provide mucoadhesiveness and pH-sensitivity properties to the co-polymer and is selected from acrylate based monomers.
mixing at least one hydrophobic drug with the said block co-polymer solution;
subjecting the resulting mixture to stirring, heating, ultrasonic treatment, solvent evaporation or dialysis to physically incorporate the hydrophobic drug into the hydrophobic core of block co-polymeric micelle, and
purifying the mixture to recover the sustained release and long residing ophthalmic formulation.
The said micelle solution of block copolymers is prepared by:
dissolving amphiphilic monomers in an aqueous medium to obtain micelles,
adding aqueous solutions of cross-linking agent, activator and initiator into the said micelles, and
subjecting the said mixture to polymerization in presence of an inert gas at 30xc2x0 C.-40xc2x0 C. till the polymerization of micelles is complete.
The said vinyl group of compounds is selected from vinyl pyrrolidone, vinyl alcohol, vinyl chloride or vinyl acetate.
Y is N-isopropyl acrylamide.
The said acrylate based monomers is selected from acrylic acid or methyl metharylate.
xe2x80x98mxe2x80x99 lies in the range 10-4000.
The said purification is done by dialysis.
The said drug is non steroidal anti inflammatory drug (NSAID).
The said NSAID drug is selected from 5-benzoyl (2,3 dihydro-1H pyrrolizone-1 carboxylic acid) (keterolac); 1-(4 chlorobenzoyl)-5 methoxy-2-methyl-1H-indole-3acetic acid (indomethacin) or N-(4-nitro-2 phenoxyphenyl) methanesulphonamide (nimesulide) or a mixture thereof or a mixture of at least any two drugs.
The said polymerized micelles containing the hydrophobic drug have a size in the range of 10 nm-100 nm diameter.
The said polymerized micelles containing the hydrophobic drug have a size in the range of 10 nm to 50 nm.
The block copolymers, made of amphiphilic monomers, are biocompatible and non-antigenic materials.
The said drug used for entrapment is in solution form or in powder form.
The solvent used for dissolving the drug is selected from dimethylformamide (DMF), dimethylsulphoxide (DMSO), dioxane, chloroform, n-hexane, dichloromethane, ethylacetate, ethanol, methanol.
The said dialysis is carried out for 2-4 hours to eliminate unreacted monomers and free NSAID, if any, at the aqueous phase.
The said drug is in hydrophobic form.
The said cross-linking agent is a bifunctional vinyl derivative.
The said bifunctional vinyl derivative is N,Nxe2x80x2 methylene bis acrylamide.
The said initiators are peroxide derivatives or 2,2xe2x80x2-azo bis isobutyronitrile (AIBN).
The said peroxide derivatives are benzoyl peroxide, or perdisulphate salts like ammonium perdisulphate.
The said activator is ferrous ammonium sulphate (FAS).
The said inert gas is nitrogen or argon.
The optimum combination of amphiphilic monomers is Vinyl pyrrolidone (VP), N-isopropyl acrylamide (NIPAAM) and acrylic acid (AA) in the molar ratio 9.5%:85.7%:4.8%.
The temperature at which polymerization is carried out ranges between 20xc2x0 C.-80xc2x0 C.
The temperature at which polymerization is carried out ranges between 30xc2x0 C.-40xc2x0 C.
The block copolymer micelles composed of polyvinylpyrrolidone (which produces hydrogel to reduce the irritability of the eye), poly-N-isopropylacrylamide (pNIPAAM) (which renders thermosensitivity) and polyacrylic acid (which renders mucoadhesiveness) to the polymeric micelles. The copolymer is having an inner hydrophobic core and an outer hydrophilic shell. To render the micellar aggregates more stable, cross-linking of the polymeric chain was done by using N,Nxe2x80x2methylene bis acrylamide during the vinyl polymerization process. The resulting micellar aggregate is suitable of dissolving hydrophobic drugs such as ketorolac, indomethacin and nimesulide. These micellar aggregates are very small in size ( less than 50 nm diameter) and therefore, can easily penetrate the mucin membrane of the eye, have mucoadhesive properties and, therefore, can adhere on the corneal surface so that the corneal penetration is an extremely slow process. At and above 35xc2x0 C., these nanoparticles are desolvated and becomes hydrophobic due to the presence of NIPAAM unit. This results the deposition of particles in the membrane pores and cause sustained release of the drug there.
The block copolymer micelles are made of mucoadhesive and thermosensitive polymer components, and when instilled, it penetrates the mucin membrane, adhere to the membrane pores and at body temperature, it becomes more hydrophobic to release the drug faster. These biodegradable block copolymer micelles nanoparticles having an average diameter of 20 nm to 60 nm (at 25xc2x0 C.) are particularly suitable for formulating an ocular delivery composition of NSAIDs like ketorolac, indomethacin and nimesulide which are usually made soluble in water by making them salts in acidic medium.
The random block copolymer of micelles of the present invention may be prepared by mixing monomers such as vinylpyrrolidone (VP), N-isopropyl is acrylamide (NIPAAM) and acrylic acid(AA) in presence of N,Nxe2x80x2 methylene bis acrylamide (MBA) and polymerizing the mixture by free radical polymerization reaction using ammnonium persulphate as catalyst. The hydrophobic moiety of the polymeric chain remain buried inside the micelles which help dissolution of drug and the hydrophilic moiety such as carboxylic acids are extended outside the surface of the micelles. The clear solution of the micellar dispersion in aqueous solution can be instilled in the patient""s eyes much more effectively and the sustained release of the drug encapsulated inside the micelles enhances the therapeutic effect of the drug.
The block copolymer used in the drug composition of the present invention may be a random copolymer of constituents VP, NIPAAM and AA.
- - - (-VP+NIPAAM+AA-)m - - - 
wherein,
m is an integer larger than 2, preferably from 10 to 4000.
In this copolymer, the VP, NIPAAM and AA components are responsible for hydrogel formation, thermosensitivity and mucoadhesiveness of the micelles respectively.
The monomers are not limited to only these three monomers only. Any monomer having these characteristics can be used for such micelles. As for example, instead of NIPAAM, one can take N-alkylacrylamide of general formula 
Wherein R1 may be a proton or CnH2n+1 in which n may have any value from 3 to 6.
Instead of VP one can take vinylalcohol.
Suitable hydrophobic drugs, which may be incorporated into the block copolymer micelles of the present invention, are non-steroid antiinflammatory drugs such as ketorolac, indomethacin and nimesulide.
The combination of amphiphilic monomers is preferably VP, NIPAAM and AA in the molar ratio 9.5%:5.7%:4.8%
In order to incorporate one or more drugs mentioned above into the block copolymer micelles, various methods described below may be used alone or in combination.
(i) Stirring
A drug is added to an aqueous solution of a block copolymer, and stirred for 2 to 24 hours to obtain micelles containing drug.
(ii) Heating
A drug and an aqueous solution of a block copolymer are mixed and stirred at 30xc2x0 C. to 80xc2x0 C. for 5 minutes to a couple of hours and then cooled to room temperature while stirring to obtain micelles containing the drug.
(iii) Ultrasonic Treatment
A mixture of a drug and an aqueous solution of a block copolymer is subjected to an ultrasonic treatment for 10 minutes to 30 minutes and then stirred at room temperature to obtain micelles containing the drug.
(iv) Solvent Evaporation
A drug is dissolved in an organic solvent such as chloroform and was added to an aqueous solution of micelles. Subsequently the organic solvent was evaporated slowly while stirring, and then filtered to remove free drug.
(v) Dialysis
The polymeric micelles solution was added to an organic solution of drug and the mixture is dialyzed against a buffer solution and then water.
The micelle solution of block copolymers is prepared by dissolving amphiphilic monomers in an aqueous medium to obtain micelles, adding aqueous solutions of cross-linking agent, activator and initiator into the said micelles, subjecting the said mixture to polymerization in presence of an inert gas at 30xc2x0 C.-40xc2x0 C. till the polymerization of micelles is complete.
The purification step is done by dialysis. The dialysis is carried out for 2-4 hours to eliminate unreacted monomers and free NSAID, if any, in the aqueous phase.
The organic solvents used for dissolving the drug is selected from dimethylformamide (DMF), dimethylsulphoxide (DMSO), dioxane, chloroform, n-hexane, dichloromethane, ethylacetate, ethanol, methanol.
The cross linking agent is a bifunctional vinyl derivative, such as N,Nxe2x80x2 methylene bis acrylamide.
The initiators are peroxide derivatives, such as benzoyl peroxide, or perdisulphate salts like ammonium perdisulphate or 2,2xe2x80x2-azo bis isobutyronitrile (AIBN).
The activator is ferrous ammonium sulphate (FAS). The inert gas is nitrogen or argon. The temperature at which polymerization is carried out ranges between 20xc2x0 C.-80xc2x0 C., particularly between 30xc2x0 C.-40xc2x0 C.
A hydrophobic drug may be incorporated into the polymeric micelles of the present invention during the time of polymerization wherein the drug is dissolved into the micelles of the monomers in aqueous solution and the polymerization is done in presence of the drug.
As the drug held in the hydrophobic core of the micelles is released on the cornea surface in a controlled manner for a long time, the composition of the present invention is suitable for formulating drugs, which are not amenable to conventional formulating techniques or using non mucoadhesive micelles.
The present invention thus provides a formulation which is therapeutically more effective and toxicologically much safer, than conventional formulations of hydrophobic drugs.